Biological reaction apparatus with draining mechanism

ABSTRACT

A biological reaction apparatus for receiving at least one substrate having a sample located in a sample region, and a separate cover, such that a reaction chamber is formed between the cover and substrate over the sample region, wherein the apparatus includes a locating means to locate the substrate; a cover locating means for locating and moving the cover with respect to the substrate; a fluid dispensing means for dispensing fluid into the reaction chamber; and a draining mechanism; wherein the draining mechanism includes wicking means.

CROSS-REFERENCE OF RELATED APPLICATIONS

This is a continuation of U.S. application Ser. No. 12/697,684 filedFeb. 1, 2010, which is a divisional of U.S. application Ser. No.12/560,850 filed Sep. 16, 2009, which is a continuation of U.S.application Ser. No. 10/518,626 filed Nov. 8, 2005, which is a nationalstage entry of PCT/AU2003/000779 filed Jun. 20, 2003, which claimspriority from Australian patent Application No. PS3114/02 filed Jun. 20,2002 and Australian patent Application No. 2003901871 filed Mar. 31,2003. The entire disclosures of each of the above-noted priorapplications are considered part of the disclosure of the accompanyingcontinuation application and are hereby incorporated by reference intheir entirety.

FIELD OF THE INVENTION

The present invention relates to a method or apparatus for providing areaction chamber for chemical reactions. The present invention alsorelates to a method of filling a reaction chamber and a fluid used forthis purpose.

BACKGROUND OF THE INVENTION

There are many applications where it is desirable to initiate a chemicalreaction on a sample. Commonly the samples are located on a microscopeslide. Typical reactions include immuno-histochemical reactions ofcellular material, or in situ-hybridisation of DNA or RNA. In otherforms, microarrays of thousands of small samples of material, includingDNA, RNA proteins or small chemical compounds are attached to amicroscope slide, where it is desirable to promote a chemical reactionbetween the material on the slide and other chemicals or fluids. Thesereactions require controlled conditions, including controlled reactiontime, temperature and concentration of chemicals. It is important thatthe reaction across the slide is uniform, and also that reactions fromslide to slide are consistent.

It is also important to minimise evaporation and overall fluid quantityused.

In the past, chemical reactions taking place on slides have beencontrolled by skilled persons adding and mixing the reagents. Thisallowed the time and quantity of the reagents to be controlled for eachslide. However, this procedure is time consuming, required highlyskilled operators, and can produce inconsistent results from slide toslide.

SUMMARY OF THE INVENTION

In one form, the present invention is a biological reaction apparatusfor receiving at least one substrate having a sample located in a sampleregion, and a separate cover, such that a reaction chamber is formedbetween the cover and substrate over the sample region, wherein theapparatus includes

-   -   a locating means to locate the substrate;    -   a cover locating means for locating and moving the cover with        respect to the substrate;    -   a fluid dispensing means for dispensing fluid into the reaction        chamber; and    -   a draining mechanism;        wherein the draining mechanism includes wicking means.

Preferably the wicking means include points of contact on the substrateto provide a fluid path to drain fluid from the substrate.

Preferably the substrates are supported in the apparatus fromunderneath. Supporting substrates from underneath removes wicking pathsfrom around the periphery of the substrate, which reduces fluid usageand loss.

In another form, the present invention provides a fill fluid forperforming a filling of a reaction chamber, where the fill fluid has aviscosity higher than an antecedent fluid on a substrate.

Preferably the fill fluid is miscible with water

Preferably the fill fluid has a higher boiling point than water.

Preferably the fill fluid leaves no residue on the substrate or sample.

Preferably the fill fluid is inert to biological reagents and samples.

Preferably the fill fluid is a solution comprising glycerol.

In one form the fill fluid contains glycerol, water, and buffer. Thebuffer may be tris buffered saline.

Preferably the fill fluid contains between 2% to 80% glycerol by volume.

More preferably still the fill fluid contains between 10%-60% glycerolper volume.

More preferably the fill fluid contains between 20% to 30% glycerol.

In one form the fill fluid includes a surfactant to aid in thedisbursement of any bubbles formed within the reaction chamber during afill cycle.

More preferably the surfactant is Tween.

In another form the present invention relates to a receptacle forsubstrates having receiving means adapted to locate a substrate and acover.

Preferably the receiving means includes stations to locate and supportthe substrate, and the cover is supported on the substrate.

Preferably the receiving stations support the substrate around part of aperiphery of the substrate.

Preferably the receiving means are defined by a respective aperturehaving peripheral ledges for supporting the substrates.

Preferably the apertures are adapted to receive support platforms from areaction apparatus, such that when loaded in a reaction apparatus, theplatforms support the substrates.

Preferably the receiving means have a lifting means for lifting thecovers from the substrate.

More preferably the lifting means are ramps adapted to engage withprojections on the cover.

Preferably the receiving means have guides allowing the cover to bemoved with respect to the receptacle and slide.

In another form the present invention relates to a dispenser for areaction apparatus including a fluid conduit,

-   -   a pump connected to the fluid conduit;    -   a locating means for moving the fluid conduit from a fluid        source to a dispensing region.

Preferably the dispenser includes a bar code sensor to detect the typeof fluid source and substrate;

Preferably the dispenser includes a means for determining the volume offluid remaining in a fluid source.

More preferably the means for determining the volume of fluid in a fluidsource includes a sensor adapted to measure the level of fluid in afluid container.

More preferably the sensor measures a change of capacitance of the fluidconduit to detect insertion into a fluid in the fluid container.

In another form the present invention relates to a method of dispensingfluid to a substrate including the steps of:

loading a reagent receptacle with at least one fluid container;

mounting the reagent receptacle to a reaction apparatus

detecting the reagent receptacle

once the reagent receptacle is detected, initiating a sensor to detectthe type of fluid within the at least one fluid container

storing the information on fluid type to allow the fluid to be dispensedonto a substrate when required.

Preferably the sensor detects bar codes.

In another form the present invention relates to a reaction apparatushaving a support projection for a slide, a dispensing means and a fluidremoval means, where the support projection is adapted to support aslide from underneath, and a wicking means contacting the periphery ofthe slide, such that the wicking means provides a wicking path to removefluid from the upper surface of the slide.

Preferably the support projection is angled between 0 and 10 degrees tothe horizontal providing the apparatus with a fluid removal region. Thisprovides a gradient to promote fluid flow.

Preferably the wicking means is wicking posts.

Preferably the wicking posts are located at the fluid removal region.

In one form the wicking means is adapted to extend across a significantproportion of the width of the substrate.

In another form the present invention relates to a reaction apparatusadapted to locate a substrate having a surface containing a sample andcover having a surface forming a reaction chamber with the samplecontaining surface, including a cover engaging means adapted to changethe volume of the reaction chamber.

This promotes mixing of fluid within the reaction chamber.

In one form the cover engaging means is a clamping mechanism adapted toclamp the cover to the substrate.

In another form the present invention relates to a reaction apparatushaving a separate substrate tray:

the substrate tray adapted to hold a number of substrates and covers;

at least one receiving station for receiving said substrate tray;

a dispensing means for dispensing fluid onto substrates in the substratetray

wherein a reaction chamber is formed between the substrate and cover,such that fluid dispensed onto the substrates enters the reactionchamber.

Preferably the reaction apparatus has a number of receiving stations,each station adapted to receive a substrate tray.

Preferably the reaction apparatus has a controller which allows thefluid to be dispensed onto a substrate on one substrate trayindependently of any other substrate tray.

In another aspect, there is provided reaction apparatus for receiving asubstrate having a sample located in a sample region and a drainingmechanism including wicking means for draining fluid from the substrate.

In another aspect, there is provided a method of forming a reactionchamber on a slide in a reaction apparatus including:

placing a cover having a cavity on a slide, forming a reaction chamber;

locating the cover and slide in a receptacle of a tray;

providing a receiving portion in the reaction apparatus having a mountfor each receptacle in the tray;

loading the tray into a receiving portion of the reaction apparatus,where the receiving portion of the reaction apparatus locates the tray;

releasably holding the cover to the slide; and

releasing the tray from the slide and cover.

In another aspect, there is provided an apparatus for loading multipleslides and covers including:

a tray having a number of receptacles for slides and covers;

a receiving portion for receiving trays;

mounts for each receptacle located in the receiving portions;

a clamp for each mount;

wherein when a tray having slides and covers is loaded into thereceiving portion, each clamp holds the cover on the slide to locate theslide, and the tray drops from the slides so each slide is supported bythe mount.

In another aspect, there is provided a method of undertaking reactionson samples on slides involving multiple steps including:

loading a first holder having at least one slide into a reactionapparatus;

scanning the slide to determine the multiple steps in the reaction totake place on the slide;

determining whether other holders have been loaded into the reactionapparatus;

undertaking the multiple steps required on the at least one slideassociated with the first holder;

when a second holder is detected, continue the steps in the reactionassociated with the at least one slides in the first holder and thenundertaking the at least one steps associated with the slides associatedwith the second holder.

In another aspect, there is provided an apparatus for performingreactions on slides including:

a tray having a plurality of receptacles adapted to support and locateslides and associated covers;

receiving ports for the trays, the receiving ports having mountsassociated with each receptacle of the tray;

a clamping mechanism for clamping the cover and slide in place;

a fluid draining means for draining fluid from the reaction chamberformed between the cover and slide;

fluid receptacles to allow at least one fluid to be placed on theapparatus;

fluid dispensing means to dispense fluid onto the slides;

wherein once the tray is loaded, the slides and cover are clamped andthe tray is moved so that the slides and covers are supported on themounts, fluid may be dispensed onto the slides by the dispensing means,and drained by the draining means.

In another aspect, there is provided an apparatus for applying reagentsto sample slides, including:

a plurality of ports for receiving the slides;

a reader for reading identification information on each of the slides;and

a reagent rack for receiving reagent containers which carry reagent tobe deposited on the slides; wherein

the slides are provided on trays, which are received in the associatedports such that each tray represents a separate batch of slides, toallow for addition and removal of separate trays, for batch processingduring operation of the apparatus.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention is described, by way of non-limiting example only, withreference to the accompanying drawings, in which:

FIG. 1 shows an example of a reaction apparatus;

FIG. 2 shows an example of a tray used with the reaction apparatus ofFIG. 1;

FIG. 3 shows the tray of FIG. 2 partially loaded into a receiving portof the reaction apparatus of FIG. 1;

FIG. 4 shows an example of a reagent container rack and rack receivingzone of the reaction apparatus;

FIG. 5 shows a robotic arm and dispensing mechanism of the reactionapparatus of FIG. 1;

FIG. 6 shows slides and covers loaded onto stations of a reactionapparatus of FIG. 1;

FIG. 7 shows a cover loaded into a tray shown in FIG. 2;

FIG. 8(a)-(c) shows a cover in three positions relative to a slide;

FIG. 9 shows a first view of an engaging means for a cover in areceiving port of the reaction apparatus of FIG. 1;

FIG. 10 shows a schematic section of a reaction chamber formed between acover and a slide;

FIG. 11 shows a washing station for the reaction apparatus;

FIG. 12 shows a station of a tray receiving port and wicking means.

FIG. 13 shows a cut away section of a cover mounted upon a slide;

FIG. 14 shows a top view of a tray receiving port of the reactionapparatus of FIG. 1;

FIG. 15 shows a cross section of a slide and cover on a mount of astation;

FIG. 16 shows a cutaway view of sections of the slide, cover, and mountof FIG. 15; and

FIG. 17 is a perspective view of a mixing station.

DETAILED DESCRIPTION

FIG. 1 shows an automated reaction apparatus 10 having bulk reagentcontainer receiving zone 12, substrate tray receiving ports 14, arobotic arm 16 and a reagent rack receiving zone 18.

Bulk container receiving zone 12 is adapted to hold a number of bulkreagent containers 20. These containers 20 typically hold fluids such astris buffered saline, PBS, Citrate, EDTA, organic solvents, wastereagents, deionised water, and dewaxing solutions. The bulk reagentcontainers of the apparatus 10 hold 1 to 4 liters of fluid.

The robotic arm 16 is moveable along the guide 24, driven by motors (notshown) and controlled by a controller (not shown) such as a computer. Asshown in FIG. 5 a dispensing means 26 is moveably mounted to arm 16, andincludes a fluid conduit such as pipette 28, for dispensing fluids. Thepipette 28 is attached by tubing 29 to a pump (not shown) which in thisexample is a motorised syringe pump capable of withdrawing, holding anddelivering an accurate volume of fluid. The pipette 28 may be loweredwhen withdrawing or dispensing fluids, and raised when moving across theapparatus 10. A sensor 33 for reading bar codes is also included on thearm 16.

The reagent rack receiving zone 18 includes 4 rack mounts 30, racklocating clip 31 and a sensor 35 for detecting the mounting of eachreagent rack 34, as best seen in FIG. 4. The reagent racks 34 eachincludes nine receptacles 36, each adapted to receive a reagentcontainer 39. The reagent racks 34 may be removed from the rackreceiving zone 18 when it is necessary to remove, refill or change acontainer 39.

In FIGS. 1 and 3 there are three slide tray receiving ports 14 and eachis adapted to hold a single slide tray 15.

The slide tray 15 (shown in FIG. 2) includes ten slide receiving means37, in the form of apertures which have support means 38. One or moresubstrates in the form of slides 1 may be placed into the slide tray 15,as shown in FIG. 3, such that the slides 1 are supported around theperiphery but not in the middle. Covers 2 are placed onto the slides 1as shown in FIG. 7. When the slide tray 15 is placed into the trayreceiving port 14, each receiving means 37 corresponds to a slidestation 35 a in the apparatus 10 as shown in FIG. 6 and described infurther detail below. A series of blocks 40 in the tray receiving ports14 are adapted to support the slides 1 when the slide tray 15 is fullyinserted into the apparatus 10 along rails 39 a. When the slide tray 15is inserted fully into the receiving port 14, it may be lowered suchthat the slides come into contact with and are supported by the blocks40. The slide tray 15 is then not in contact with the slides, leavingthe slides supported from underneath by the blocks 40. While only twoslides 1 and covers 2 are shown loaded onto the tray 15 shown in FIG. 3,there may be any number of slides and covers, up to the number ofreceiving means 37 contained by slide tray 15.

The blocks 40, which are typically metal and may be controllably heatedor cooled, support the slides 1 in conjunction with wicking means 41 inthe form of wicking posts 42 as shown in FIG. 12. The upper surface ofblocks 40 are inclined at a small angle to the horizontal (typically 5degrees) to promote fluid flow along the slide during operation of theapparatus 10.

The cover 2 (best seen in FIGS. 8 and 13) is one of a number ofvariations possible, other variations being described in copendingInternational patent application titled “A cover for a slide” by thesame applicant and hereby incorporated by reference. The cover 2 is madefrom a clear plastic material, and is substantially the same width asthe slide 1 to which it is to be mounted. A cavity 51 is located on sidea of the cover 2 that faces the sample, and this cavity 51 inconjunction with lands 52 and sample holding surface 53 of the slideforms a reaction chamber 32 as shown in schematic FIG. 10, where the zaxis has been exaggerated for clarity. FIG. 10 is a sectioned view of acover over a slide 1 showing the reaction chamber 32, sample 5, lands 52and slide surface 53. Typically the slide is 25 mm wide by 76 mm long,and the cavity is 100 micrometers high. The land 52 is in closeproximity to or contacts slide surface 53 along contact surface 54 asshown in FIG. 13, and therefore restricts fluid leakage from thereaction chamber 32 outside the reaction chamber. Capillary forcesassist in holding the fluid in the reaction chamber 32.

A locator arm 3 enables the cover 2 to be moved along the slide 1 by alocator engaging means 43 shown in FIG. 9. Each locator arm 3 is engagedby a bracket 44. A range of positions of the cover relative to theslides is shown in FIG. 8, where FIG. 8 (a) is fully open, FIG. 8(b) ispartially open and FIG. 8(c) is fully closed. A reaction chamber 32 isformed between the cover 2 and slide 1 over a sample 5 on the slide 1when the cover is in a closed or partially open position. The cover 2includes a fluid reservoir 19 where fluid may be dispensed. There areseveral forms of fluid reservoir, as described in the abovementionedcopending application. The cover and slide are capable of holding fluidin the reservoir 19. when the cover is in contact with the slide.

The fluid in the reservoir is drawn into the cavity 51 of the cover asthe cover moves over the slide from an open position shown in FIG. 8(a)to a closed position shown in FIG. 8(c). The reservoir 19 may holdsufficient volume such that there is still fluid in the reservoir whenthe cover is in a closed position, and this provides a reservoir offluid to reduce the need for fluid top ups during extended reactiontimes or sustained high temperatures. It is believed that the fluid isdrawn into the cavity by a number of factors including capillary forces.

The covers 2 include wings 50 projecting from cover 2 adapted to engageramps 52 on the slide tray 15, as shown in FIG. 7. The wings lift thecover 2 clear from the slide 1 when the wings 50 on the cover 2 engagelifting means in the form of ramps 52. It is possible to move the cover2 to a position where the sample is uncovered but the cover remains incontact with the slide, along guides 56. Depending on the configurationof the ramps 52 and wings 50, it may not be necessary to completely openthe chamber before the cover loses contact with the slide 1.

The arm 3 is moved by an actuator such as a cam arrangement (not shown)which engages positioning member 45 controllably so that the cover isable to be accurately positioned with respect to the slide along thex-axis shown in FIG. 8. While FIG. 9 shows that all covers are moved atonce, in other examples of reaction apparatus it is possible to haveindividual control of the covers by moving arms individually.

In FIG. 6, slides 1 having bar codes 6 are shown on their respectiveblocks 40. For the purposes of this diagram the slide tray 15 andengaging means 43 have been omitted from view for clarity. A clamp 60 isused to hold a cover 2 securely in position on the slide 1 during aprocessing step. Clamp 60 includes a number of legs 62, which aresituated around the periphery of the slide 1 and have spring likeproperties to provide an even force around the periphery of the cover.The clamp 60 may be made from a plastic material, and in another example(not shown) the legs may be made from metal, in the form of a spring(leaf or coil). Other forms of legs or clamp are possible such ascompressible foam or pneumatic clamps.

The clamp 60 for each cover 2 may be raised when the cover 2 is to bemoved, or lowered to engage the cover 2 during a fluid dispensingoperation. In the present example, all clamps 60 and covers 2 in aparticular receiving port 14 are moved together. Individual receivingports 14 may operate independently of each other.

In use, bulk reagents in bulk reagent containers 20 are loaded into theapparatus 10. Reagent racks 34 having reagent containers 39 are loadedinto the rack mounts 30. Sensors 35 detect their presence and the barcode sensor 33 reads the bar codes on each reagent container 39 toidentify the contents of each reagent container 39 relative to itsposition in the reagent rack 34. Information relating bar codes 6 onslides 1 to samples on the slides 1 and bar codes 6 on reagentcontainers 39 relating to their respective contents, is input into thecontroller (not shown), which is typically a computer work stationhaving an appropriate software interface and drivers. A slide tray 15containing at least one slide 1, but up to ten slides, is placed intothe receiving port 14, whereupon a sensor (not shown) detects the slidetray 15 and initiates a scan of the stations 35 a. When scanning, thebar code sensor 33 on the robotic arm 16 moves to each station 35 a andattempts to read a bar code 6. If a slide 1 with a bar code 6 ispresent, the controller compares the bar code 6 with a list of knownslides and information input by the user to determine which protocol toapply to each individual slide 1. Alternatively, once the bar codes havebeen scanned, the user inputs information required for the apparatus toprocess the slide. Each slide may have a different protocol. Thecontroller compares the reagents required to perform the reactionsdictated by the protocols with the reagents located in the containers 39in the reagent racks 34. Any discrepancy will cause an error message tobe sent to the user. If a reagent container 39 is missing then thereagent rack 34 may be removed and the correct container 39 placed inthe rack 34, whereupon the rack 34 is detected and another scan ofreagent containers 39 is undertaken.

If no errors are present, the robotic arm 16 moves the pipette 28 of thedispensing means 26 to the appropriate reagent container 39 andwithdraws the required amount of fluid. At this time the dispensingmeans 26 checks the capacitance of the pipette 28, which changes whenthe pipette comes into contact with the fluid surface of a reagentcontainer 39. In this way the volume of fluid remaining in the reagentcontainer 39 can be determined and the user can replace the container 39as necessary. The robotic arm then moves the pipette 28 to a first slide1 (determined by the controller) and dispenses the fluid onto thesurface of the slide 1. There are several options in placement of thepipette 28 and cover 2 in relation to the sample 5 on the slide 1, andthese will be discussed further below.

Once the dispensing operation for a first slide 1 has been undertaken,the process is repeated for further slides. It is not necessary for eachslide 1 to be filled with the same fluid at each step, and the slidesmay be filled in any order that is appropriate. A washing station 120shown in FIG. 11 is located near the reagent racks 34 and may be used toclean the pipette 28 prior to withdrawal of a different reagent. Washingstation 120 includes a receptacle 121 for receiving the pipette 28,where cleaning fluid from one of the bulk reagent containers 20 ispumped onto the outside of the pipette 28 to remove traces of theprevious fluid. Cleaning fluid may also be pumped from the bulk reagentcontainer 20 via tubing to clean the inside surfaces of the pipette 28.

Reagents may be pumped from the bulk reagent containers 20 throughpiping and valves (not shown) into the pipette 28. Bulk reagent from thebulk reagent containers 20 may also by pumped to a wash station 120.

Other reagent containers such as the bulk reagent containers 20,included in the body 12 of the apparatus 10, can add to the type ofreagents that may be dispensed onto the slide. Some bulk reagentcontainers 20 normally contain fluids required for washing and hydratingsamples.

The reagent rack 34 may be used to contain a detection kit. A detectionkit consists of a number of reagents in separate reagent containers 39that are used to perform a particular test on one or more samples. Sucha detection kit may include nine reagent containers 39 to perform asingle test, and this reduces the number of reagent containers 39available to other slides to twenty seven.

Typical reagents applied to samples on slides include primaryantibodies, such as those sold by Novocastra Laboratories Ltd. Thesereagents are normally supplied in the reagent containers 39 in volumestypically between 7 ml and 30 ml. Other reagents and fluids, such asbuffers and de-ionised water, may be kept in the bulk storage containers20 which typically have volumes between 1-4 liters.

Some reagents, once prepared for application to a sample, have arelatively short shelf life. Therefore, either the reagent is suppliedpre-mixed in a ready-to-use formulation, whereupon it must be usedwithin a short period of time from ordering, or it may be prepared bylaboratory staff prior to use, and placed into an appropriate reagentcontainer. Some of the reagents, such as 3′,3-diamino benzidene (DAB),when in a final form, begin to degrade soon after preparing and may notbe useable more than 24 hours after initial preparation. This requires anew batch to be prepared every day, and ensuring that old batches arediscarded after use. Further, enzymes such as protease may need to beapplied in varying concentrations depending on factors such as tissuetype, other reagents to be applied etc. This can result in numerousbatches of reagents being required to be prepared before application tothe samples, with the associated problems such as correct application,expiry date, correct mixing, tracking and traceability.

Concentrated primary antibodies may also require preparation before use,requiring dilution before application to a sample. Primary antibodiescan be supplied either in a concentrated form or pre-dilutedready-to-use. However, it may be necessary to have several differentworking dilutions of the same antibody on a single apparatus 10, whichwould otherwise take up several locations in the reagent rack 34. It istherefore advantageous to have a single reagent container 39 of anantibody, where diluting of the antibody reagent may take place beforethe reagent is applied to the sample. The primary antibody may bediluted by a primary antibody diluent such as ABDIL 9352 sold by VisionBioSystems Ltd.

In the present embodiment of the apparatus 10, a mixing station 122 isprovided, as shown in FIG. 11. Mixing station 122 includes an insert130, as shown in FIG. 17, having a number of mixing vials 132. Theinsert 130 has six vials, each vial able to hold a different reagent.The vials 132 are shown all the same volume, but may vary in volumeaccording to requirements. Typical volumes may be 7 ml per vial.

Also mounted to the insert 130 is a tab 134. Tab 134 may be used toidentify the insert 130 such as by way of a barcode. It is envisagedthat as the insert 130 is disposable, but may contain a number ofdifferent reagents over the course of several runs of the apparatus 10.

The bar code on the insert 130 may be used to identify the insert 130 sothat the controller knows when to discard the insert 130, and requestthat a new insert be loaded into the mixing station 122. This may bepredetermined after a set period of time or uses.

Also shown on insert 130 is an overflow aperture 135, which is adaptedto allow excess fluid to drain from the insert should any of the vials132 overflow.

In use, information from the slide bar codes may be cross-checked with adatabase in the controller to establish which series of reagents is tobe applied to each slide. The apparatus 10 then compares the reagentsrequired, to the reagents currently loaded. If a reagent is identifiedthat is not in final form for application to a sample, then a preparingstep is scheduled into the order of tasks to be undertaken on theapparatus 10.

In one example, three reagent containers (identical to reagent container39 located in the reagent rack 34) each have a component part A, B, andC of DAB may be located on the apparatus 10. In the present example DABwill be mixed in a ratio of 1 part A to 25 part B to 1 part C. To mix abatch of DAB ready for use, the robotic arm 16 first moves to thereagent container containing part A, and withdraws a set volume of partA of the reagent. The robotic arm 16 then moves to one of the vials 132at the mixing station 122 and deposits the volume into one of the vials132. The pipette 28 then moves to a washing station 120 located next tothe mixing station 122, where the outside and inside of the pipette 28are rinsed. Once cleaned, the robotic arm 16 moves the pipette 28 to thereagent container containing part B of the reagent. The pipette 28withdraws the reagent (25 times the volume of part A) and moves to thevial containing part A. Once deposited in the vial, the pipette 28 movesto the washing station and is again washed, before moving to the reagentcontainer holding part C of the reagent. The same volume as removed fromthe container holding part A is removed, and the pipette 28 moves to theoriginal vial and deposits the reagent with the other reagents.Initially depositing the reagents into the mixing vials causes somemixing, however additional mixing can be accomplished by withdrawingsome or all of the reagent from the vial 132 into the pipette 28, thenre-depositing the reagent into the vial 132. The pipette 28 may movevertically to ensure that the tip is above the fluid level whendepositing to aid the mixing process. The energy of re-deposition causesthe reagents to mix more readily. This mixing process can be undertakena number of times as desired. After the reagent has been mixedsufficiently, the pipette 28 may proceed to the wash station 120 if thenext reagent to be applied to a sample is not DAB. This volume of thevials and the amount withdrawn by the pipette 28 provide a sufficientvolume of DAB for many applications to samples. Whenever DAB isrequired, the robotic arm 16 moves the pipette 28 to the vial where theDAB was mixed, as the vial in which mixing of particular reagents isrecorded by the controller. The time of the preparation is alsorecorded, so that after a predetermined period of time the mixed reagentcan be discarded. This prevents the prepared reagent from being usedafter expiring.

After completion of testing for the day, or at the expiry of the DAB,the vial 132 containing the DAB (or any other reagent that has expired)can be cleaned as discussed below.

In relation to scheduling of mixing within a batch, specific details ofscheduling are disclosed in Australian Provisional patent applicationtitled “Method of Scheduling” filed 24 Feb. 2003 by same applicant, thecontents of which are hereby incorporated by reference.

While the above process is automated, the resources employed (roboticarm 16 and pipette 28) may be utilised for significant periods of timein general reagent application to samples, and therefore it may bedesirable to reduce the necessity to prepare several batches of reagentduring a day. For this reason the apparatus 10 can be programmed toprepare reagents in the absence of any samples loaded into the apparatus10 or during normal processing, and the volume and concentrations areuser determinable through a user interface (not shown).

In the above example the concentration and time of preparation of eachreagent in each vial 132 are stored in the memory of the controller ofthe apparatus 10, so there is no chance of old or incorrect mixedreagent being applied to a sample, reducing operator error.

The mixing by the pipette 28 ensures that the prepared reagent is fullymixed before application to a sample, and provides a better uniformityof mixing than, for example, applying components of the reagent directlyto the sample and mixing on the sample.

Other examples of reagents that benefit from mixing on the apparatus 10include protease, which may be required to be applied in a number ofconcentrations. In the above example, only one reagent container ofprotease would be required, and several concentrations of protease maybe prepared by the apparatus 10 using diluent stored on board either ina reagent container 39 or bulk reagent container 20. These differentconcentrations may be placed in different vials 132 for later use.

In the above example, it is possible to have the mixing tasks scheduledinto the steps of applying reagent to the samples. For example, thereare often periods of time during a testing of a slide where there are notasks required of the robot arm. These times may be referred to as opentimes, which typically occur when the fluid applied to a slide requirestime to react before the next step is undertaken. If an open time is ofa sufficient length, it may be possible to schedule in a mixing step.This minimises the time required to complete the application of fluid tosamples, while freeing the operator from preparing the reagents.

After reagent is prepared, and it is applied to samples, remaining orexpired prepared reagent is siphoned to waste by the aspirator. Thevials 132 may then be cleaned. Cleaning is undertaken by draining anyprepared reagent remaining after the required prepared reagent has beendispensed. Draining is done with the pipette 28, the drained fluid beingdirected to an internally plumbed bulk waste container. Oncesubstantially empty, a rinse cycle is undertaken. The rinse cycle mayuse a cleaning solution, which for example could contain an alcohol suchas IMS dispensed into the vial 132. The cleaning solution is thendrained via the pipette 28. More than one rinse cycle may be undertaken.After removing cleaning solution for the final rinse, any remainingcleaning solution is allowed to evaporate to completely empty the vial.

It is also possible to revisit the mixing vial after a predeterminedtime from initial preparation, to re-mix the reagent. This may be doneby withdrawing some of the prepared reagent into the pipette 28, andredispensing into the same vial 132. This may be important wherecomponents of the prepared reagent settle after time or do not staymixed after a period of time. As with initial mixing, the remixing stepmay be scheduled during a period of inactivity of the robot arm and anaspirator.

When a slide tray 15 is loaded into the apparatus, each brackett 44 isengaging the locator arm 3 of each cover 2 in the slide tray 15. If anopen fill is required, ie where the cover 2 is substantially or fullywithdrawn from the slide 1, the locator engaging means 43 moves allcovers 2 on the slide tray 15 off the slides to a position such as thatshown by cover 2 in FIG. 8(a). This open position of the cover 2 exposesthe sample 5, whereupon the pipette 28 may be positioned in a variety ofpositions. The positions of the pipette 28 include either over thesample 5, to dispense fluid directly onto the pipette 28, or adjacentthe front of the cover 2 into a fluid reservoir 19 shown in FIG. 8. Thereasons for each position will be explained below.

In an open fill situation, once the fluid has been dispensed on allslides, the locator engaging means 43 moves to position the reactionchambers 32 over the samples on the slides. Capillary action and themovement of the cover 2 over the surface of the slide 1 causes dispensedfluid to flow into the region between the cover 2 and slide 1. The clamp60 may be used to hold the cover 2 in place and prevent it from floatingon the film of liquid between the cover 2 and slide 1.

When the slide 1 is on the block 40, it may be in contact with wickingposts 42, as shown in FIGS. 14 and 15. Movement of the slide 1 on theblock 40 is possible as slide lengths vary, and movement of the cover 2over the slide can move the slide 1. Normally this movement is only inthe order of 1-2 mm. In another example (not shown) it is possible touse an actuator to move the slide away from the wicking posts to reducewicking of fluid from the reaction chamber.

FIG. 15 shows the cover 2 on the slide 1, both located on block 40. Thewicking posts 42 are in contact with the slide and therefore provide awicking path for fluid. The reaction chamber is located between theslide and cover but as FIG. 15 is approximately to scale, it cannot beclearly seen in this view. Fluid entered in fluid reservoir 19 flowsinto the reaction chamber and may flow from the reaction chamber downdrain 55 associated with the wicking posts 42. To assist in fluidclearance, the air pressure around the wicking posts may be lowered bywithdrawing air from the drain 55 by a pump such as a fan (not shown).This will promote fluid flow through the reaction chamber and out thedrain 55 if required. Withdrawing the cover from the slide will alsopromote fluid flow down the drain 55.

The wicking posts will wick fluid even if not touching the slide, as themeniscus of the fluid will extend out from the edge of the slide nearthe wicking posts if there is fluid pressure from the wicking posts, orif the air pressure in that region is reduced.

The wicking action may, however, be interrupted if required, such asduring an incubation period, by manipulating the locator arm 3 so as tomove the cover 2 away from the wicking posts 42 a distance sufficient toprevent any further drain of fluid from the reaction chamber.

When dispensed fluid fills the reaction chamber 32 there may be fluidcontact between the fluid in the reaction chamber and the wicking posts42. The upper surfaces of the blocks 40 are at angles approximately 5degrees to the horizontal with the end of the slide adjacent the wickingposts lower than the bar code end of the slide. The angle promotes fluidflow towards the wicking posts 42, which provide the only contact withthe slide 1 apart from the block 40. As the wicking posts 42 contact theslide 1 at or near the upper surface of the slide 1, at the lowest endof the slides upper surface, the fluid will tend to wick from the areain the reaction chamber on the slide adjacent the wicking posts 42 andnot from other areas, as there are no other wicking points.

It is possible to control the dispenser 26 to dispense fluid onto theslide in various positions. The fluid may be dispensed towards the bardcoded end of the slide, or towards the wicking post end of the slide ifthe cover is in an open position. It is also possible for the dispenserto dispense in a “staggered waterfall” arrangement where fluid isdispensed in a number of positions up the slide. The cover may close asthe dispenser moves up the slide.

Fluid is dispensed onto the slide 1 in controlled volumes. It has beenfound that in the current arrangement, fluid does not wick from thereaction chamber 32 down the wicking posts 42 unless one of twoconditions are met. Firstly, there needs to be fluid in the reservoir 19to push fluid through the reaction chamber 32. The additional fluiddisplaces the antecedent fluid, which is removed from the reactionchamber. The antecedent fluid is removed from the reaction chamber viathe wicking posts. Thus it is possible to replace a fluid in thereaction chamber by placing fluid in the fluid reservoir. Secondly, apump can produce a reduced atmospheric pressure around the wicking poststo cause the pressure differential to draw fluid from the reactionchamber. The reaction chamber may also be drained by reducing airpressure around the wicking posts.

If no new fluid is to be added to the reaction chamber it is possible todrain the reaction chamber by opening the reaction chamber. This isaccomplished by sliding the cover along the slide 1 until the sample isuncovered. The fluid in the reaction chamber will tend to follow thecover off the sample, draining the fluid via the wicking posts.Alternatively, it is possible to turn on the fan to draw fluid from thereaction chamber, where the cover can remain in a closed position. Acombination of the above is possible.

In some cases, such as where the fluid being applied or in the reactionchamber is particularly viscous, it may be necessary to utilise the pumpand apply fluid to the reservoir to cause fluid flow through thereaction chamber. In this way it is possible to change over fluid acontrolled way.

The cover 2 and slide 1 are removed from the apparatus 10 when thereaction is complete and therefore the reaction chamber 32 is unique toeach reaction. This eliminates the necessity to thoroughly clean astatic reaction chamber as required in other apparatus. Further, thereaction chamber is substantially sealed to the environment reducingevaporation and the possibility of the sample drying out.

As the reaction chamber is formed from a slide and a replaceable cover,it is relatively inexpensive to form a reaction chamber, and a new,clean reaction chamber is formed for each reaction, reducing cleaningcosts and time, as well as eliminating the possibility of crosscontamination with previous reactions or cleaning fluids.

The initial fill with the cover withdrawn (open fill) provides a methodof filling the reaction chamber while minimising the formation of voidsor bubbles inside the chamber. Due to the reaction chamber having adepth of approximately 100 microns, once the cover is over the slideforming the reaction chamber, it is difficult to flush the chamber ofbubbles or voids. Some of the fluids used in the reactions are extremelyexpensive and may be hazardous, and therefore it is desirable to keeptheir consumption to a minimum.

A suitable initial fill fluid has been found to be a mixture of waterand 25 to 30% glycerol. Small amounts of glycerol do assist in reducingthe incidence of bubble formation, as do larger amounts, however it hasbeen found that in some circumstances 25% glycerol by volume works well.Additives such as detergents (Tween for example) may be included toreduce surface tension, which also have proved beneficial in removingvoids in some circumstances.

The use of glycerol reduces the propensity of the fluid to wick from thesurface of the slide via extraneous wicking paths. This reduces thenumber of large voids that form during an initial fill.

To assist in removing any voids that may reside in the reaction chamberafter an initial fill, it has been found that a fluid having reducedsurface tension and viscosity, but miscible with water, such as analcohol like isopropanol, is useful as a flushing fluid.

Typically flushing occurs after a heating phase, as increasing thetemperature in the reaction chamber can cause bubbles or voids to form.The use of a low viscosity fluid such as isopropanol can assist inmoving the bubbles or voids.

Once the reaction chamber is filled with fluid, it is possible to addfurther fluid without entrapping additional air. Thus, it is possible tochange fluids by merely topping up the fluid reservoir, and in someinstances, reducing air pressure near the wicking posts. The reactionchamber thus formed exhibits some desirable flow characteristics, inthat a new fluid will not tend to mix with the fluid it is replacing.The capillary nature of the reaction chamber does not allow significantturbulent mixing and therefore it is possible to accurately time thechanging of fluids without requiring extensive flushing of the chamberor slide surfaces. This allows the start and finish of a reaction to bedetermined with sufficient accuracy across a range of reactions andfluids.

The speed of the cover movement and pressure reduction can effect thevolume of residual fluids left behind.

In order to promote reactions in the reaction chamber on the sample, itis possible to move the cover vertically (in the z axis direction asshown in FIG. 8) on the slide by modulating the load on the clamp 60.The vertical movement assists in mixing the fluid in a verticaldirection as well as a direction across the slide (y-axis direction),rather than along its length. Filling and draining the reaction chambermove fluid along the length of the slide (x-axis direction) and this maybe assisted by moving the cover along the x-axis of the slide by movingthe arm 44. The blocks 40 may be heated to promote the reaction.

It is desirable in many reactions, for example involving in-situhybridisation, epitope retrieval, or dewaxing, to heat the fluid in thereaction chamber to a temperature approaching 100 degrees Celsius. Inthis situation, gas bubbles have been known to form, and the gas bubblescan be difficult to shift. If the bubbles occur on the sample theyreduce the amount of fluid exposed to the sample, and can thereforeeffect the consistency of the result within a sample, as well as betweensamples on different slides. In such situations it has been found thatusing covers having one or more coatings can reduce the incidence ofbubble formation.

Another feature of the reaction apparatus 10 is that the size of thereaction chamber may be varied. Typically the volume of the reactionchamber when the cover is completely over the slide, termed the closedposition, is 150 microliters. However, if the cover is not completelyclosed then the reaction chamber formed between the cover and slide maybe of reduced volume. In FIG. 8(b) a cover in a partially closedposition is shown, wherein the volume of the reaction chamber would besignificantly reduces, for example to 80 microliters. This example maybe useful where samples are small, or placed towards an end of the slidthat allows the cover to form a smaller reaction chamber while stillcovering the sample. Smaller reaction chambers require smaller volumesof fluids, which is advantageous if the fluids used are expensive ordifficult to obtain. The examples of the reaction apparatus allow theposition of the cover to be referenced when dispensing fluid onto theslide. Therefore, when the cover is in the open position, it is possibleto dispense fluid either on top of the tissue sample, or between thetissue sample and the cover, so that movement of the cover to a closedposition pushes fluid across the sample while filling the reactionchamber. It is also possible to dispense fluid at a number of positionsalong the slide, or to dispense fluid on or near the front edge of thecover.

The following is a description of set up and use of the above-describedapparatus.

1. Slide Loading:

Paraffin-embedded tissue sections (sample 5) mounted onto glass slidesare loaded into the slide tray 15 with covers 2 and inserted into thereceiving zones 14 of the reaction apparatus 10. The user selectsdesired protocols, run type [ie 100 μL (economy—⅔ of slide) or 150 μL(standard-full slide)] and ensures that the reagents trays 34 containingthe necessary reagent containers 39 are loaded into the apparatus 10.

2. Dewaxing:

Removal of wax from tissue sections following sectioning is requiredprior to performing staining procedures. For dewaxing on the instrumentthe cover remains in a closed position while dewaxing solution isdispensed by the dispensing means 26 onto the slides, which arepre-heated to 70° C. by mounting blocks 40. Slides are incubated for 4min at 70° C. prior to removal of excess dewaxing solution by reducedair pressure around the wicking posts caused by a pump (not shown).Fresh dewaxing solution is dispensed onto the slides for incubation at70° C. for a further 4 min. This process is typically repeated once morefor all slides in a tray that require dewaxing. Slides are cooled toambient temperature and covers opened and closed to remove excessdewaxing solution containing residual dissolved wax. All slides arewashed with isopropanol applied by the dispensing means one slide at atime, to remove remaining dewaxing solution, and then all slides arerehydrated with distilled water dispensed by the dispensing means.

3. Epitope Retrieval:

Before IHC and ISH processing can take place, it is necessary to exposeepitopes (proteins, DNA, RNA) within the tissue which may have becomehidden during the fixation process. On the instrument two protocols maybe present:

a. Heat-Induced Epitope Retrieval (HIER)

Following dewaxing, all slides receive an initial fill of retrievalbuffer (initial fill fluid) (10 mM Sodium Citrate/30% Glycerol/0.05%Tween) with the cover in the open position to facilitate movement ofsolution down the slide and reduce bubble formation. Covers are closedand mounting blocks 40 heat the slides to 100° C. for the requiredretrieval time. After retrieval is finished, slides are cooled byindividual flushing with retrieval buffer by the dispensing means.

b. Enzyme-Induced Epitope Retrieval (EIER)

Protease solution (ie proteinase K, pepsin, and trypsin) is dispensedonto each slide by the dispensing means and incubated for 10-30 minutesat the desired retrieval temperature (for example ambient-50° C. or roomtemperature). After retrieval is complete, each slide is washed withdistilled water dispensed by the dispensing means.

4. Immunohisochemistry (IHC):

IHC is based on specific binding of antibodies (proteins) to antigens(proteins) in tissue biopsies and specimens. Following the epitoperetrieval stage, each slide receives buffer containing Tween-20 from thedispensing means. Each slide may be treated with hydrogen peroxide for 8min at ambient temperature to block endogenous peroxidase activitywithin the tissue sections and is washed with TWB buffer containingTween-20, again dispensed by the dispensing means. A primary antibodydirected against a specific target protein is applied by the dispensingmeans to the tissue sample and incubated for 15-60 min. This is followedby a secondary biotin-labelled antibody incubation. Bound antibody isdetected by dispensing streptavidin- or alkaline phosphatase-conjugatedperoxidase onto each slide, which is visualised by addition of achromogen (ie DAB, BCIP/NBT), all by dispensed by the dispensing means.Sections are counterstained with hematoxylin, also dispensed by thedispensing means.

5. In Situ Hybridisation (ISH):

ISH allows the detection of specific nucleic acid sequences within acell. Following the EIER stage, tissue sections are dehydrated bydispensing isopropanol into the reaction chambers of each slide and thecover moved to the open position to dry the tissue. A fluorescein- orbiotin-labelled nucleic acid probe is applied to the slide and the coverclosed slowly to distribute the probe evenly across the tissue. Theprobe is allowed to hybridise to its complementary DNA/RNA target in atissue section for 1.5-2 hours at 37-55° C. Where the target is DNA, thetissue section and probe are first denatured at high temperature (ie 95°C.) for 5-10 min prior to hybridisation. Slides are washed by dispensingTWB from the dispensing means using a staggered waterfall rinse togently remove unbound probe. Following washing, the cover is moved tothe closed position for the remainder of the procedure. Bound probe isdetected by applying an anti-fluorescein or anti-biotin antibodyconjugated to alkaline phosphatase, dispensed from the dispensing means,which is visualised by addition of an enzyme substrate (BCIP/NBT), alsodispensed from the dispensing means.

6. Removal:

Once the protocol has been completed for a particular slide tray, thetray may be removed regardless of the status of the other slide trays.As the slide tray may contain slides each having different protocolsapplied, the tray must remain in the apparatus until all protocols forthat particular tray have been completed. An indicator such as a lightinforms the user when all the protocols to be applied to the slides onthe slide tray have been completed.

Once the reaction chamber has been filled it is possible to hold thesample in a buffer for an extended period of time. Fluid in the reactionchamber can be topped up if, for example, some slides reactions arecompleted but other slides on a slide tray require additionalprocessing. Having three slide trays allows a certain amount offlexibility in that samples that require time intensive processing canbe placed in one slide tray, while faster processing may be undertakenon a separate slide tray. An additional slide tray may be entered whileone or more slide trays have begun processing, and it is possible toremove a finished slide tray while another slide tray is beingprocessed. The reagent racks 34 may be removed during a process run, iffor example, a container empties. Once the reagent rack 34 is replaced,the bar code sensor 33 scans the bar codes on the reagent containersagain to ensure that only the correct reagents are applied.

The dispensing mechanism employs a sensor to detect the level of thefluid in the reagent container, and therefore warns the user when thecontainer is running low. This is important as reagent may have a shortuseful life when not stored properly, and the reagent is also expensive,therefore there are significant advantages in reducing waste.

The sensor may be attached to the pipette to sense when the pipettereaches the surface of the fluid in the reagent container. This allowsthe volume of a container to be determined, and a warning maybe sent tothe operator is fluid levels drop to a predetermined level. The reagentrack may then be removed from the apparatus, the container replaced,whereupon the scanner will determine whether the correct reagent wasreplaced by reading the bar code on the reagent container. In this wayoperator error is reduced.

There are a number of variations described herein, but the apparatus isdesigned to allow a flexible approach to fluid application, reactiontime and temperature. It is therefore not intended that the apparatus belimited to particular examples of potential methodology, as variationsin fluid application, cover position and movement.

The protocols that may be applied are varied, and it is possible toapply a different protocol to each sample on a slide in a single rack.Further, it is possible to load a new tray of slides or remove acompleted tray of slides while the apparatus is processing another trayof slides.

Without limiting the forgoing, some specific aspects of the inventionare recited below, together with a brief description of some advantagesof each:

A method of forming a reaction chamber on a slide in a reactionapparatus including:

placing a cover having a cavity on a slide, forming a reaction chamber;

locating the cover and slide in a receptacle of a tray;

providing a receiving portion in the reaction apparatus having a mountfor each receptacle in the tray;

loading the tray into a receiving portion of the reaction apparatus,where the receiving portion of the reaction apparatus locates the tray;

releasably holding the cover to the slide; and

releasing the tray from the slide and cover.

The above-mentioned method allows a slide and cover to be easily placedinto receptacles in a tray. The tray may have a number of receptacles,for example 10 receptacles per tray as shown in the figures of theembodiments disclosed herein. The tray can then be loaded into areceiving portion of the reaction apparatus, so that, for example up to10 reaction chambers formed from slides and covers, can be placed intothe reaction apparatus. As the tray is located by the reaction apparatusupon loading, and the slides and covers are located by the tray, theexact position of up to 10 reaction chambers can be determined easilywithin the apparatus. Given that slide dimensions vary due tomanufacturing inaccuracies, and the covers do not contact the sides ofthe slides (to eliminate extraneous wicking points), such that thecovers can move freely, on top of the slides if not constrained by othermeans, locating 10 reaction chambers at once can be difficult.

Once the tray is loaded and the slides and covers are fixed in positionby the clamps, the tray can be removed. In the present examples the trayis dropped down so that the mounts support the slides and covers. Thisremoves all contact around the edges of the slides except for thewicking posts. Thus it is possible with this arrangement to easily andquickly locate a number of slides and covers without any contact withthe sides of the slides. As the covers do not have a positive sealingarrangement, and the reaction chamber is generally full of fluid, thisarrangement assists in loading multiple slides without fluid lossthereby minimising bubble formation within the reaction chamber.

An apparatus for loading multiple slides and covers including a trayhaving a number of receptacles for slides and covers;

a receiving portion for receiving trays;

mounts for each receptacle located in the receiving portions;

a clamp for each mount;

wherein when a tray having slides and covers is loaded into thereceiving portion, each clamp holds the cover on the slide to locate theslide, and the tray drops from the slides so each slide is supported bythe mount.

Preferably, a draining means is provided.

Preferably the draining means includes a wicking means.

The apparatus above allows slides and covers to be loaded easily by anoperator, in batches if required.

A method of undertaking reactions on samples on slides involvingmultiple steps including:

loading a first holder having at least one slide into a reactionapparatus;

scanning the slide to determine the multiple steps in the reaction totake place on the slide;

determining whether other holders have been loaded into the reactionapparatus;

undertaking the multiple steps required on the at least one slideassociated with the first holder;

when the second batch is detected, continue the steps in the reactionassociated with the at least one slides in the first holder and thenundertaking the at least one steps associated with the slides associatedwith the second holder.

This is possible in some situations as there are usually gaps where theapparatus used to start or stop reactions, or undertake other tasks(such as the pipette mounted to the robot arm) are not utilised all thetime.

If the apparatus are used all the time then the steps of the reaction totake place on the at least one slides associated with the second holderwill not commence until the first bath has finished.

An apparatus for performing reactions on slides including a tray havinga plurality of receptacles adapted to support and locate slides andassociated covers

receiving ports for the trays, the receiving ports having mountsassociated with each receptacle of the tray;

a clamping mechanism for clamping the cover and slide in place;

a fluid draining means for draining fluid from the reaction chamberformed between the cover and slide;

fluid receptacles to allow at least one fluid to be placed on theapparatus

fluid dispensing means to dispense fluid onto the slides

wherein once the tray is loaded, the slides and cover are clamped andthe tray is moved so that the slides and covers are supported on themounts, fluid may be dispensed onto the slides by the dispensing means,and drained by the draining means.

Preferably, the apparatus includes a locating means for locating andmoving the cover with respect to the slide.

Preferably there is a locating means associated with every receptacle ina tray.

Preferably all locating means associated with a particular tray all moveat the same time to move the cover with respect to the slide, tofacilitate fluid dispensation or draining of all slides on a tray.

An apparatus for applying reagents to sample slides, including:

a plurality of ports for receiving the slides;

a reader for reading identification information on each of the slides;and

a reagent rack for receiving reagent containers which carry reagent tobe deposited on the slides; wherein

the slides are provided on trays, which are received in the associatedports such that each tray represents a separate batch of slides, toallow for addition and removal of separate trays, for batch processingduring operation of the apparatus.

The batch loading, again, provides substantial flexibility for anoperator insofar as testing and scheduling is concerned.

The invention claimed is:
 1. An apparatus comprising: a controller; arobotic arm; a dispensing mechanism, mounted on the robotic arm,configured to dispense fluids; a plurality of individual tray receivingports disposed under the robotic arm; a plurality of slide trays, eachof the plurality of slide trays comprises a sliding mechanism such thatsaid respective single slide tray is fitted to slide into a respectiveindividual tray receiving port of the plurality of individual trayreceiving ports, wherein each slide tray comprises a plurality of slidereceiving sections; a plurality of slides, a single slide of theplurality of slides placed on each slide receiving section of theplurality of slide receiving sections of each slide tray; a plurality ofcovers, each cover mounted on a slide of the plurality of slides,wherein each cover comprises a cavity on a side facing the correspondingslide thereby forming a reaction chamber between each slide and itscorresponding cover; and a rack receiving zone comprising a reagent rackwhich includes a plurality of reagent containers, wherein the controllercomprises non-transient memory including a program to control thedispensing mechanism of the robotic arm to dispense fluid onto a slideof one of the slide trays independently of dispensing fluid onto anotherslide of another one of the slide trays, wherein the sliding mechanismis rails that extend along a longitudinal side of the sliding tray,wherein each of the plurality of individual tray receiving ports arehousing the respective single slide tray of the plurality of slidetrays, wherein the plurality of slide trays are fitted to slide into therespective individual tray receiving ports via the respective rails,wherein the rails are on both sides of each of the plurality of slidetrays and slide along the longitudinal sides of the respective trayreceiving ports, and wherein, in a fully inserted position of therespective single slide tray into the slide receiving port, the slidetray is not in direct contact with the slides in the slide receivingsections and the slides come in contact and are supported, fromunderneath, by a plurality of blocks of the respective individual trayreceiving port, wherein the plurality of blocks support respectiveslides of the plurality of slides.
 2. The apparatus of claim 1, whereinthe cover comprises an arm portion which slides the cover along the topof the slide thereby changing a size of the reaction chamber in responseto the sliding of the arm portion.
 3. The apparatus of claim 2, whereineach of the slide trays comprises a pair of ramps including slots, andthe cover comprises wings which engage the pair of ramps in the slots inresponse to sliding of the arm portion.
 4. The apparatus of claim 3,further comprising: an actuator which engages a positioning member tomove the arm portion to slide the cover over the slide, wherein theactuator is controlled by the controller.
 5. The apparatus of claim 3,further comprising: a clamp which secures the cover to the slide duringprocessing of a sample on the slide.
 6. The apparatus of claim 1,wherein each of the tray receiving ports comprises the plurality ofblocks corresponding to the plurality of slide receiving sections in therespective slide trays, wherein the plurality of blocks support theplurality of the slides.
 7. The apparatus of claim 6, wherein when therespective slide trays are received in the respective tray receivingports, the plurality of blocks support the plurality of the slides andthe respective slide trays are not in direct contact with the pluralityof slides.
 8. The apparatus of claim 6, further comprising: a pluralityof wicking posts respectively provided for the plurality of the blocks,wherein the plurality of wicking posts further supports the plurality ofthe slides.
 9. The apparatus of claim 8, wherein a block, from among theplurality of the blocks, comprises an upper surface inclined towards awicking post, from among the plurality of the wicking posts, providedfor the block, wherein the upper surface is inclined with respect to ahorizontal of the slide tray.
 10. The apparatus of claim 1, wherein thecontroller is programmed to remove and replace the reagent rack with anew reagent rack without interrupting operations of the robotic armduring a process run in which the robotic arm is in use.
 11. Theapparatus of claim 1, wherein the dispensing mechanism comprises a fluidconduit which dispenses fluids, wherein a plurality of samples forundergoing reactions are respectively placed on the plurality of slides,and in response to at least one of the tray receiving ports receivingthe respective slide trays, the controller is programmed to scan theplurality of slides to determine a reaction to be performed at eachindividual slide.
 12. The apparatus of claim 11, wherein based on thedetermined reaction, the controller is programmed to perform adispensing operation by determining a reagent required to perform thereaction at the slide, and determine whether at least one of theplurality of reagent containers includes the determined reagent.
 13. Theapparatus of claim 12, wherein in response to determining that at leastone of the plurality of reagent containers includes the determinedreagent, the controller is programmed to continue the dispensingoperation by controlling the robotic arm (i) to move the dispensingmechanism to the at least one reagent container to withdraw thedetermined reagent from the at least one reagent container into thefluid conduit, (ii) to move the dispensing mechanism to the slide, and(iii) to dispense the withdrawn reagent from the fluid conduit into thereaction chamber.
 14. The apparatus of claim 13, wherein the controlleris programmed to repeat the dispensing operation for each individualslide on the respective slide trays to process the slides as a batch.15. The apparatus of claim 14, further comprising: a washing stationcomprising: a receptacle for receiving the fluid conduit, wherein thecontroller is programmed to pump cleaning fluid from a bulk reagentcontainer into the receptacle to clean the fluid conduit betweenperformances of the dispensing operation for each individual slide. 16.The apparatus of claim 12, further comprising: a bulk reagent containersection; and a plurality of bulk reagent containers stored in the bulkreagent container section, wherein in response to determining that atleast one of the plurality of reagent containers does not include thedetermined reagent, the controller is further programmed to continue thedispensing operation by controlling the robotic arm (i) to determinewhether at least one of the bulk reagent containers includes thedetermined reagent and (ii) to refill, in response to determining thatat least one of the bulk reagent containers includes the determinedreagent, the at least one reagent container in the reagent rack with thereagent included in the at least one bulk reagent container, (iii) tomove the dispensing mechanism to the at least one reagent container towithdraw the determined reagent from the at least one reagent containerinto the fluid conduit, (iv) to move the dispensing mechanism to theslide, and (v) to dispense the withdrawn reagent from the fluid conduitinto the reaction chamber.
 17. The apparatus of claim 1, wherein therails of each of the plurality of slide trays are positioned below theplurality of slide receiving sections and wherein the rails are on bothsides of the plurality of slide trays.
 18. The apparatus of claim 1,further comprising: a locator arm which moves the corresponding coverrelative to the respective slide from among the slides via a locatorengaging element, wherein the locator arm is engaged by a bracket fromamong a plurality of brackets corresponding to the plurality of slidereceiving sections, and wherein each of the plurality of individualreceiving ports operate independently of each other.
 19. An apparatuscomprising: a controller; a robotic arm; a dispensing mechanism, mountedon the robotic arm, configured to dispense fluids; a plurality ofindividual tray receiving ports disposed under the robotic arm; aplurality of slide trays, each of the plurality of slide trays comprisesa sliding mechanism such that said respective single slide tray isfitted to slide into a respective individual tray receiving port of theplurality of individual tray receiving ports, wherein each slide traycomprises a plurality of slide receiving sections; a plurality ofslides, a single slide of the plurality of slides placed on each slidereceiving section of the plurality of slide receiving sections of eachslide tray; a plurality of covers, each cover mounted on a slide of theplurality of slides, wherein each cover comprises a cavity on a sidefacing the corresponding slide thereby forming a reaction chamberbetween each slide and its corresponding cover; and a rack receivingzone comprising a reagent rack which includes a plurality of reagentcontainers, wherein the controller comprises non-transient memoryincluding a program to control the dispensing mechanism of the roboticarm to dispense fluid onto a slide of one of the slide traysindependently of dispensing fluid onto another slide of another one ofthe slide trays, wherein the sliding mechanism is rails that extendalong a longitudinal side of the sliding tray, wherein each of theplurality of individual tray receiving ports are housing the respectivesingle slide tray of the plurality of slide trays, wherein each of thetray receiving ports comprises a plurality of blocks corresponding tothe plurality of slide receiving sections in the respective slide trays,wherein the plurality of blocks support the plurality of the slides,wherein the slides come in contact and are supported, from underneath,by the plurality of blocks of the respective individual tray receivingport, and wherein when the respective slide trays are received in therespective tray receiving ports, the plurality of blocks support theplurality of the slides and the respective slide trays are not in directcontact with respective slides of the plurality of slides.
 20. Theapparatus of claim 19, wherein the cover comprises an arm portion whichslides the cover along the top of the slide thereby changing a size ofthe reaction chamber in response to the sliding of the arm portion. 21.The apparatus of claim 20, wherein each of the slide trays comprises apair of ramps including slots, and the cover comprises wings whichengage the pair of ramps in the slots in response to sliding of the armportion.